Difference between revisions of "Part:BBa K1036001"
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<b>Construciton</b><br/> | <b>Construciton</b><br/> | ||
− | <img src="https://static.igem.org/mediawiki/2013/6/67/XMU-China_ndh-fusion-pcr.png" width= | + | <img src="https://static.igem.org/mediawiki/2013/6/67/XMU-China_ndh-fusion-pcr.png" width=800px; height=300px;/><br/> |
<b>SDS-PAGE</b><br/> | <b>SDS-PAGE</b><br/> |
Revision as of 05:14, 27 September 2013
the ndh gene coding for respiratory NADH dehydrogenase II
Description
ndh is the coding gene for NADH dehydrogenase II (NCBI), a non-proton-translating single subunit membrane bond enzyme important in maintaining a redox-balanced dinucleotide pool. NADH dehydrogenase II is the preponderant respiratory source of H2O2 as well as O2-. In fact, substantial H2O2 was formed when quinoneless vesicles containing only NADH dehydrogenase II were incubated with NADH.
Earlier studies estimated that the formation rate of intracellular O2- was about 10-5 M S-1 by extrapolation of O2- yield from this enzyme. And NADH dehydrogenase II formed about 4 O2- per 10 molecules of H2O2 both during respiration and in the quinoneless membranes.
Experimental Data
Construciton
![](https://static.igem.org/mediawiki/2013/6/67/XMU-China_ndh-fusion-pcr.png)
SDS-PAGE
To test the expression of ndh gene, an experiment was designed. The strain E. coli BL21(wide type) carried plasmid pSB4K5-ndh was cultured in LB medium with 25 μg/ml kanamycin under the condition of 37oC,200 rpm for 12 hours. Then the cultured medium was added into 100 volumes of flesh LB medium with 25 μg/ml kanamycin and cultured under the same condition. 10-3 mM AHL was added into the medium after 2 hours and continued culturing for more 10 hours. The control was without AHL. The cell was harvested, washed twice by ddH2O and ran in SDS-PAGE.
The result was showed in Fig.1 and ndh means without AHL and ndh+AHL means with AHL. The red and blue arrows showed the two up-regulation proteins, while the green arrow showed the down-regulation protein.According to the LC-MS-MS data, the band showed by the red arrow was the ndh gene.
![](https://static.igem.org/mediawiki/2013/2/2e/XMU-China_SDS-PAGE-ndh.png)
Sequence and Features
Barcodes are discontinued, but one was appended to the sequence of this part. Composite parts using this part will include the barcode. More ...
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1360
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]